外泌体载药系统在PD模型中跨血脑屏障递药的多模态成像研究

Cumulating evidence has emerged for the function of exosomes as ideal central nervous system drug delivery vehicles, since they can easily overcome blood brain barrier. Besides, exosomes are key mediators of information transmission between cells and are well tolerated in the body as evidenced by their ubiquitous distribution in biological fluids. However, the delivery pathway and distribution in vivo, metabolism process and therapy mechanism of the drug loaded exosomes is still unclear and needs to be clarified. In this regard, the development of non-invasive, in situ, real-time and dynamic imaging analysis method for monitoring drug delivery process of exosome is the key of studying the pharmacology of exosome-drug complex, looking for exosome targets, as well as investigating the therapy mechanism. Herein, we develop multifunctional nanoprobes integrating magnetic resonance imaging and optical imaging agents with different properties to allow precise, high contrast, high resolution imaging of exosomes in excessively activated microglia induced Parkinson’s disease (PD) rat modal. The multifunctional nanoprobes labeled exosomes could be robust analysis method for intracellular tracking, screening the optimal exosome candidates for targeting delivery suppressing over activated microglia drug into lesion and facilitate the establishment of versatile therapeutic tools for PD ultimately. Our results may put forward new approaches to PD treatment by vesicle transport, which has significant clinical applications and social values.

外泌体来源于机体自然成分，能穿透血脑屏障，具有良好的转运能力及体液中稳定存在的特点，显示出其在中枢神经系统药物载体方面的巨大优势。然而，载药外泌体在体内代谢机制不明，严重限制其向临床转化的进程。建立无创、原位、实时、动态监测载药外泌体跨血脑屏障、靶向病灶过程的成像分析方法是监测外泌体分布，寻找作用靶点以及探索作用机理的关键。本项目以小胶质细胞过度激活的帕金森病(PD)动物模型为研究对象，构建具有高分辨率、高对比度和高灵敏度的磁共振和光学多模态成像纳米探针，采用多模态成像技术观察外泌体载药系统脑内分布和药物释放情况，阐明其在PD治疗中的作用及机制。项目开发了一个跨血脑屏障递药的新技术，通过探讨其可行性及技术操作性，为今后发展外泌体载药治疗模式奠定理论基础及实验基础，具有重要的临床应用价值与社会意义。

外泌体来源于机体自然成分，能穿透血脑屏障，具有良好的转运能力及体液中稳定存在的特点，显示出其在中枢神经系统药物载体方面的巨大优势。然而，载药外泌体在体内代谢机制不明，严重限制其向临床转化的进程。建立无创、原位、实时、动态监测载药外泌体的成像分析方法是监测外泌体分布，寻找作用靶点以及探索作用机理的关键。本项目构建了CD63适配体功能化的量子点（QDs-Apt），特异性标记干细胞源外泌体。通过凝胶电泳和荧光光谱仪验证了QDs上成功偶连适配体。QDs-Apt与梯度离心纯化制得的脐带间充质干细胞源外泌体共孵育后提纯，荧光标记的Exo分别与人肝细胞QSG-7701、人肾小管上皮细胞HKC和人正常肺上皮细胞BEAS-2B共孵育，发现Exo进入三种细胞的排序为QSG-7701>BEAS-2B>HKC，表明Exo与肝脏细胞的亲和性最强。研究中发现，外泌体的产率低，限制其用作药物载体。因此，本项目开发了一种将单细胞悬液反复挤压通过不同粒径的聚碳酸酯膜制备细胞膜纳米囊泡的方法，该方法产率高，操作简便，质量可控且可重复性高。通过超声处理的方法，使细胞膜纳米囊泡负载姜黄素（NV-Cur）。NV-Cur作用于MPP+诱导大鼠原代神经元损伤PD模型，展现了良好的神经元保护作用。